Influenza A viruses (IAV) are significant human pathogens causing yearly epidemics and occasional pandemics. Past pandemics have resulted in significant morbidity and mortality. The 1918 influenza pandemic was thought to have resulted in the death of at least 675,000 people in the U.S., and 40 million people worldwide. Pandemics in 1957 and 1968, while less severe, were also of major public health importance. A novel influenza A virus of swine origin became pandemic in 2009, causing the first pandemic in 41 years. In addition, annual epidemic influenza causes are also very significant resulting in up to 49,000 deaths in the US annually. Highly pathogenic avian H5N1 viruses continue to circulate enzootically in poultry in many countries and continue to cause human infections. Recently a novel avian H7N9 strain emerged in China also causing human infections and fatalities. A variety of experimental pathogenesis studies to model host adaptation, map viral virulence factors and host factors in disease progression, evaluate novel therapeutics and vaccines were performed. Understanding the origin, virulence, and pathogenic properties of the 1918 virus, is crucial for current public health preparedness and future pandemic planning. The origin of the 1918 pandemic virus has not been resolved, but its coding sequences are very like those of avian influenza virus. Recent work with 1918:low pathogenicity avian influenza H1N1 chimeric viruses suggested that the virulence factor(s) in the 1918 hemagglutinin (HA) gene might be shared with low pathogenicity avian influenza viruses expressing H1 HAs. Work is ongoing to assess experimental mouse pathogenicity with low pathogenicity avian influenza A viruses with all 16 subtypes. These viruses are also serving as challenge strains in experiments designed to evaluate influenza viral-like particle (VLP) vaccines. Previous work showed that a neuraminidase N1 VLP vaccine, derived from the 2009 pandemic H1N1 virus, could protect mice given a lethal highly pathogenic H5N1 virus challenge. In follow-up studies done in collaboration with Dr. Maryna Eichelberger at the FDA, neuraminidase (NA) immunity was further studied. Twelve NA amino acids essential for MAb binding to the NA of a recent seasonal H1N1 virus were mapped using monoclonal antibodies (mAbs). Several of these residues were recognized by strain-specific mAbs, while several other residues were within conserved epitope(s), allowing cross-reactive mAbs to bind the NAs of seasonal H1N1 and the 1918 and 2009 pandemic (09pdm) H1N1 as well as H5N1 viruses. A single dose of mAbs administered prophylactically fully protected mice against lethal challenge with seasonal and 09pdm H1N1 viruses and resulted in significant protection against the highly pathogenic wild-type H5N1 virus. The identification of conserved N1 epitopes reveals a molecular basis for NA-mediated immunity between H1N1 and H5N1 viruses and demonstrates the potential for developing broadly protective NA-specific antibody treatments for influenza. Influenza A virus infection leads to variable and imperfectly understood pathogenicity. We recently reported that segment 3 of the virus contains a second open reading frame (X-ORF), accessed via ribosomal frameshifting. The frameshift product, termed PA-X, comprises the endonuclease domain of the viral PA protein with a C-terminal domain encoded by the X-ORF and functions to repress cellular gene expression. PA-X also modulates IAV virulence in a mouse infection model, acting to decrease pathogenicity. Loss of PA-X expression leads to changes in the kinetics of the global host response, which notably includes increases in inflammatory, apoptotic, and T lymphocyte-signaling pathways. Studies on the role of PA-X in influenza virus replication and pathogenesis are ongoing. In a collaboration with Dr. Paul Digard at the University of Edinburgh, another novel protein product of influenza A viruses was identified, in segment 7 of the virus. Serial passage of an influenza virus lacking the M2 mRNA2 splice donor site identified a single nucleotide pseudoreverting mutation, which restored growth in cell culture and virulence in mice by upregulating segment 7 mRNA4 synthesis rather than by reinstating mRNA2 production. The mRNA4 was shown to encode a novel M2-related protein (designated M42) with an antigenically distinct ectodomain that can functionally replace M2.